Process for the treatment of hyperuremia of nephritis

ABSTRACT

Process for the treatment of hyperuremia of nephritis comprising administering to a patient 2-30 grams per 24 hours of sarcosyluric acid.

United States Patent Francis Albert Kirsch 64, Rue de Prony, 75 Paris, France [2]] Appl. No. 713,802

[72] Inventor [22] Filed Mar. 18, 1968 [45] Patented Sept. 21, 1971 [3 2] Priority Mar. 20, I967 [33] France [31] 99,444

[54] PROCESS FOR THE TREATMENT OF HYPERUREMIA 0F NEPHRITIS 1 Claim, 1 Drawing Fig.

[52] US. Cl 424/253 [51] Int. Cl A6lk 27/00 [50] Field of Search 424/253 [56] References Cited OTHER REFERENCES Chemical Abstracts 7 I: 128689 q 1969) Mylius, Bcr., 1884, 17, pages 517 to 527 Primary Examiner.ler0me D. Goldberg Attorney-Young & Thompson ABSTRACT: Process for the treatment of hyperuremia of nephritis comprising administering to a patient 2-30 grams per 24 hours of sarcosyluric acid.

PATENTEU SEPZI i971 3,509.080

AWm me PROCESS FOR THE TREATMENT OF HYPERUREMIA OF NEPHRITIS The present invention relates to a process for the treatment of hyperuremia of nephritis.

This composition is particularly remarkable in that it contains sarcosyluric acid as active principle.

Sarcosyluric acid is known chemical compound the therapeutical properties of which had not been evidenced heretofore.

It was described, in particular, by Mylius (Ber. 1884, 17, 517) under the name of sarcosinuric acid. Mylius described also a process for the preparation of this compound, comprising reacting uric acid with excess sarcosine above the melting point (2 l 2 C.) of the latter.

Sarcosyluric acid has the following physical-chemical properties:

1t decomposes without melting from a temperature of 300 C. According to its elemental analysis, it has the following empirical formula:

c,H,,N,0,-2n,o.

Calculated, for this formula: C 34.91 4.76 N 25.45%

found: 7 35.17 4 .68 25.53

Paper (Whatman Nl) chromatography in propanol-water-ammonia 2:721 system shows a sitlgle U V. spotofRfQSO (ascending method).

Sarcosyluric acid is more water-soluble than uric acid, in an amount of about 17 g./liter at 20 C. Solubility increases considerably at alkaline pH values. An ammonium salt is readily prepared by dissolution in dilute ammonia solution followed by concentration in vacuo. This salt and similar salts may also be used, just as sarcosyluric acid, as active principle of the therapeutical composition according to the present invention.

The results of elemental analyses conform with those obtained by Mylius: sarcosyluric acid crystallizes with two molecules of water; this water may be lost on extended heating in vacuo at about 100 C.

Spectrographic investigations provided the following data:

1. the UV spectrum of sarcosyluric acid determined in water shows a strong shift of the bands present in the spectrum of uric acid. This shift may correspond to a suppression of the resonance effects caused by the introduction of sarcosine. ln water, uric acid shows three bands at 203,233 and 287 mu with a shoulder at about 295 my; sarcosyluric acid shows three bands at 200, 216 and 259 mp with a shoulder at 265 2. The IR spectrum was determined with a suspension in pure paraffin oil. It possesses a highly complex set of bands. It-

was compared with the spectrum of uric acid. One finds particularly the presence of a strong acute band at 2.8g. which may be assigned unambiguously to the NH of sarcosine.

3. The NMR spectrum determined in D in a medium made alkaline with K CO is rather odd. A single signal is present, a singlet at 3.06 p.p.m. assigned to the methyl protons at the NH of sarcosine. No signal is visible for the CH of sarcosine: this peculiarity is assigned to proton exchange with deuteriums, which protons are located on a carbon a to the amide (exchange facilitated by the alkaline pH).

4. The mass spectrum determined using a ms9 apparatus at 280 shows the expected molecular ion at m./e. 239, together with an abundant ion at m./e. 168 due to the loss of (M 71). By its mass, the ion at 168 corresponds to uric acid. The mass spectrographic behavior of sarcosyluric acid is consistent with the contemplated structure of a fixation of sarcosine by the carboxyl.

Taking such data into account, it follows that sarcosine is attached to uric acid by its carboxyl group, the site of this binding being impossible to determine with certainty. It appears likely, however, that this binding occurs at the NH group at 1- position of uric acid, which is apparently the more highly activated.

Thus, the most likely structural formula of sarcosyluric acid The pharmacological investigation carried out with sarcosyluric acid demonstrated the following data:

l-Oral Subacute Toxicity in Mice.

A lot of 20 male mice, having an average weight of 18-20 g., was given orally, by means of a probang, l g./kg. of sarcosyluric acid 5 days per week, during 3 weeks. The product was suspended in a 5 percent gum syrup solution. A lot of 10 male mice was used as reference and was given only gum syrup.

The weight curve of the treated animals is found to be identical with that of the controls.

No fatal issue and no sign of toxicity were noted during the time of treatment. At the end of the treatment, 10 treated mice were sacrificed and autopsied, the other 10 being kept under observation during 8 days.

Visible macroscopic injuries could be found in neither of the lots on examination of the organs.

llShort Run Oral Toxicity in Rats A lot of 24 Wistar rats, 12 male and 12 female, weighing -130 g. were given orally, by means ofa probang, l g./kg. of sarcosyluric acid 5 days per week, during 11 weeks. The product was suspended in a 5% gum syrup solution. A lot of 24 rats, 12 male and 12 female, was used as reference and was given only gum syrup.

In the treated lot, lmale rat and 1 female rat died at the beginning of the treatment (at the end of the second week). No visible macroscopic injuries of the organs could be detected on autopsy.

The weight curve of the treated animals (see Tables 1 and 2 below) is:

in the case of the female rats, identical with that of the reference animals;

in the case of the male rats: the weight gain is slightly higher than that of the controls, from the 7th week on, and until the end of the treatment.

Uric acid determinations were carried out at the end of the experimentation, both in the treated rats and in the controls. After taking blood samples at the level of the carotid, the determinations were carried out in serum, according to the uricase method.

As is apparent from the data set forth in Tables 1 and 2, the average uric acid level in the treated lot is lower than that of the reference lot. The difference is significant for p=0.05.

No macroscopically visible injury was noted on autopsy.

TABLE 1 MALE RATS Reference Animals Treated Animals Table l Continued Average TABLE 2 FEMALE RATS Reference Animals Treated Animals In addition to the safe character of sarcosyluric acid, its activity on the normal metabolism of uric acid and, hence, of nucleoproteins, development is evidenced by such pharmacological tests.

The following tests were carried out for the purpose of fully demonstrating the decrease of blood urea level on administration of sarcosyluric acid:

EXPERIMENTAL PROCEDURE Two reference female rabbits having an average weight of 3 kg. were given intravenously l ml./kg. of a aqueous arginine hydrochloride solution (i.e., I g./kg. of arginine hydrochloride). Blood samples were taken: 5 minutes prior to injection of arginine, than 1 hour, 2 hours, 3 hours, 4 hours, 6 hours, 8 hours and 24 hours after injection. Serum urea determination was carried out with each sample using a specific micromethod involving urease.

A lot of 8 female rabbits having an average weight of 2.5 kg. was given orally I00 mg./kg. of sarcosyluric acid as a 1.5% solution in distilled water: I g./kg. of arginine was then injected intravenously:

to 3 rabbits after administration of sarcosyluric acid;

to 3 rabbits 1 hour after administration of sarcosyluric acid;

to 2 rabbits 24 hours after administration of sarcosyluric acid.

Blood samples were taken: 5 minutes prior to injection of arginine, and then I hour, 2 hours, 3 hours, 4 hours, 8 hours, 24 hours and 30 hours after this injection. Serum urea was determined in each sample thus taken.

\\'ITII RESPECT TO UREA LEVEL PRIOR TO THE IN- JECTIOX OF ARGININE Treated with sarcosyluric acid 100 n1g./kg.

Time alter Controls 30 minutes 24 hours artrininc, (argininc prior to 1 hour prior prior to hours (i.v.) alone) arginine to argininc arginiuo I 24 25. 8 I8. 4 .29. I 2 35 41, .5 24. 5 42. 5 3 78. 5 43. 2 33. 4 41. 2 4 75 43. 7 34. 7 41. 2 3i). 7 21. 5 28. 4 50. 1 11. 4 38. 5 2 r 35. 8 2. 5

In the accompanying drawing the single FIGURE illustrates such results by means ofa graph showing the percent increase of the urea level as a function of time. Curve A relates to the controls who were given only I g./kg. (i.v.) of arginine; curve B relates to the animals given I00 mg./kg. of sarcosyluric acid orally 30 minutes prior to the administration of argininc; curve C relates to the animals given I00 mg./kg. orally, one hour prior to the administration of arginine, and curve D relates to the animals given 100 mg./kg. orally, 24 hours prior to the administration of arginine.

Thus, the therapeutical composition according to the invention may be used:

in the treatment of disorders of the metabolism of uric acid and of nucleoproteins, particularly in the treatment of gout in which sarcosyluric acid may be associated with an anti-inflammatory drug such as aspirin or phenyl butazone.

Moreover, it is apparent from the above tests that sarcosyluric acid opposes ureogenesis and is capable either of reducing hyperureogenesis or of inhibiting exaggerated catabolisms.

In may also oppose neoglucogenesis in diabetics in whom a hyperglycemia peak may be noted after an injection of arginine.

This action would result from deamination inhibition, a mechanism which appears to be involved against the neoformation of glucose, especially when a fasting or diabetic organism is concerned.

In such various applications, the drug may be administered by the oral, parenteral and rectal routes, the active principle being associated with the vehicles and excipients suitable for such various routes of administration.

Thus, suitable although nonlimiting pharmaceutical forms are tablets, perfusion vials and suppositories.

By the oral or rectal routes, 2-6 g. daily of active principle are administrable, the unit dosage forms of the drug containing 250-500 mg. of active principle.

On perfusion, 530 g. daily of active principle will be advantageously administered.

For the purpose of additionally illustrating the invention, six case reports in which the drug according to the invention (SUA) was administered as tablets containing 500 mg. of active principle are given below.

CASE REPORT 1 Mr. MAI... E., 52 years old gardener. Typical gout since the age of 40. Iophus of the big toe for the past three months. Regularly treated with COLCHICINE I mg. and ASPIRIN l g. Is given 4 tablets daily of SUA, during 1 week, while continuing to take both ASPIRIN and COLCHICINE.

Good tolerance.

Prior to SUA: uricemia=l03 mg. p.l.

After SUA: uricemia =99 and 77 mg. p.l.

CASE REPORT 2 Mr. VAY... R., 60 years old. Onset of disease at the age of 39. Substantial tophaceous chronic polyarticular gout since several years. Renal lithiasis. Regularly treated with 1 mg. of COLCHICINE. Is given 4 tablets daily of SUA during 1 week,

' while Continuing to take COLCHICINE.

Good tolerance.

Prior to SUA: uricernia= I IS mg. p.l.

uricuria 875 mg. per day. After SUA: uricemia 90 and I00 mg. p.l.

uricuria =540 mg./24 hrs. CASE REPORT 3 Mr. DAV... F., 45 years old. First attack at the age of42. No tophus. Treated for gouty polyarthritis with COLCHICINE 2 mg. per day. Is given 4 tablets daily of SUA, during 1 week, while continuing to take COLCHICINE.

Good tolerance.

Prior to SUA: uricemia 80 and 56 mg./l.

uricuria 380 and 640 mg./day After SUA: uricemia= 52 rng./l.

uricuria 350 mg./day

Urea, glycemia, blood protides with SA/SG, total and esterified cholesterol were determined and the functional hepatic tests were carried out, prior to and after SUA: all these tests remained normal.

CASE REPORT 4 Mr. MAI..., RJ., 67 years old. First attack at the age of 32. Small tophi. Admitted to hospital for gouty polyarthritis treated and cured with COLCHICINE.

Treatment used prior to and concurrently with SUA: IN- DOMETHACINE I mg./day, COLCHICINE 1 mg. ULTRA LEVURE (Saccharomyces culture preparation). Was given SUA at the rate of I tablet on the Ist day, 2 tablets the 2nd day, 3 tablets the 3rd day, then 4 tablets per day, total time of treatment with SUA being 7 days. On the 6th day appeared hydarthrosis of the knee, not very painful.

Good tolerance.

Prior to SUA: uricemia 86, 96, 86 and 92 mg./I. uricuria 560, 800 and 450 mgJday Alter SUA: uricemia 98 mg./l.

uricuria 500 mgjday Blood urea, glycemia, blood protides with SA/SG, total and esterified cholesterol, urinary sugar and albumin were determined prior to and after SUA: all such tests remained normal.

CASE REPORT Mr. ROU... R., 38 years old. Gout since the age of 35. No tophus. Admitted to hospital for sciatica. During hospital stay, moderate gouty fluxion of both big toes. Is given 4 tablets daily of SUA during 1 week. During treatment, the fluxion of both toes heals gradually and completely.

depressive neurosis in 1960,

chronic ethylism.

Clinical condition during hospitalization; attack of gout which is improving under the influence of a treatment combining COLCHICINE I mg. ASPIRIN 3 g., SUPPO NEMBUTAL I, IDOMETACINE 100 mg.

Is given 4 tablets daily of SUA, during 1 week, together with the above-described treatment. During SUA treatment: no new attack of gout.

Tolerance: some nausea, with headaches, during the 2 first days of treatment with SUA.

Prior IO SUA uricemia I08 mg.ll.

uricuria 470 mgjday uricemia 85 and 70 mg./l.

uricuria 478 and 380 I mg./24 hrs.

After SUA:

Clearance of urea was determined under SUA =77% and 83%. Total protides, SA/SG, hepatic tests determined prior to and after SUA remained normal.

When several uricemia ad uricuria values are given After SUA," the first value was obtained on the last day of treatment, (the patient taking the drug) the others were obtained I-3 days later.

When a single value is given, the sample was taken on the last day of treatment.

The following remarks can be made on inspection of the case histories set forth above:

a. TOLERANCE SUA was generally well tolerated.

The biological tests carried out, relating in particular to Qfunctioning of the liver, functioning of the kidneys, to

protides, showed no anomaly attributable to SUA. b. ACTION ON URICEMIA This action is summarized in Table 4 below:

TABLE 4 Original uricemia less uricemia Original uricemia less uricemia on Table 4 shows that, under treatment, uricemia decreased four times form 4 to 25 mg.; it remained stationary in case report 5 and increased by 8 mg. in case report 4. The average decrease of uricemia, under treatment, is I0 mg.

In the 4 case reports where uricemia was determined l-3 days after the end of treatment, it exhibited a decrease of 14-38 mg. with respect to initial uricemia, i.e., an average decrease of 23 mg.

c. ACTION ON 24-HOUR URICURIA In the four cases where uricuria was determined prior to and after treatment, it was found to decrease during treatment (Case report 2: 425; Case report 3: Case report 4: 103; Case report 6: 45). Average decrease of uricuria is 183 mg.

d. ACTION ON URIC ACID CLEARANCE In case report 5, uric acid clearance measured prior to and after treatment with SUA was unchanged.

To conclude, it follows from said case reports that the drug according to the invention is endowed with a reducing action on uricemia, an action which does not take place at the level of the kidneys.

Having now described my invention what I claim as new and desire to secure by Letters Patent is:

1. Process for the treatment of a patient suffering from hyperuremia of nephritis wherein said patient is administered 2-30 grams per 24 hours of sarcosyluric acid. 

